The central focus of this project is to test the hypothesis that the Ifitm proteins down-modulate cell activation and prevent viral infections by shuttling modifying enzymes to the cell surface to degrade activation complexes. The activation of cells of the immune response often requires signaling through cell surface receptors. This event brings into activation domains, such as lipid rafts, both the activating proteins (kinases, etc) and the substrates (receptors and intracellular signaling partners). In some cells (phagocytic cells such as macrophages and B cells) many such complexes are internalized and degraded. Other activation complexes on these and other cells, however, remain on the cell surface and require modification to block further cellular activation. The Ifitm proteins have been characterized as functioning to help down-regulate activated cells, depressing adhesion and proliferation. In addition they were recently characterized as the major determinants of the anti-viral state in mammalian cells after activation with type I and type II interferons. However, the mechanism by which the Ifitm proteins accomplish these effects has not been described. We have found that the Ifitm proteins can bind to a variety of proteins whose functions are associated with the modification of proteins by the addition of substrates (such as ubiquitin or SUMO) or enzymatic cleavage. We propose that the Ifitm proteins are held within the cell and are released following activation to intercalate within the membrane. As the Ifitm proteins make this migration, they carry these modifying proteins as cargo for association with proteins within the membrane. The delivery of these cargo proteins into activation complexes is followed by their action to degrade and/or modify surface proteins thus depressing cell activation. This same pathway is also proposed to function to control viral infection. We propose to target one specific cargo protein in particular, Bat5, which has been shown to bind to the Ifitm proteins of man and mouse. We further propose that the absence of this pathway will allow for uninhibited cellular activation potentially leading to increased cell proliferation and lack of immune control that could lead to autoimmunity and tissue damage. PUBLIC HEALTH RELEVANCE: The control of cell activation requires pathways that down regulate activating complexes and components. The Ifitm proteins have been implicated as helping depress cell activation and enhance innate immune defenses to certain viral infections but no mechanisms as to how they accomplish this have been demonstrated. We propose to test the hypothesis that the Ifitm proteins shuttle cargo enzymes into membrane activation complexes. These enzymes then target constituents of such sites for modification and degradation thus depressing activation and inhibiting infection.